Transcription in vitro directed by a mutant form of DNA-dependent RNA polymerase II from a mouse lymphoblastoid cell line will be analyzed. The mutant RNA polymerase is resistant to the toxin alpha-amanitin. Differences between alpha-amanitin-resistant and wild-type RNA polymerase with respect to DNA binding sites, rates of elongation, RNA chain length, temperature dependence of initiation, stability of initiation complex, and sensitivity to rifamycin AF/O13 will be investigated. Both chromatin and protein-free DNA will serve as template. Results will be interpreted in terms of structure-function relationships of RNA polymerase. In addition, temperature-sensitive RNA polymerase mutants in a Chinese hamster cell line will be isolated. Mutants will be selected on the basis of survival of killing due to (1) the incorporation of radioactive nucleosides and (2) incorporation of the toxic nucleoside analogs tubercidin and 6-thioguanine.